The purification of proteins for the production of biological or pharmaceutical products from various source materials involves a number of procedures. Therapeutic proteins may be obtained from plasma or tissue extracts, for example, or may be produced by recombinant methods such as by cultures of eukaryotic or prokaryotic cells containing at least one recombinant plasmid encoding the desired protein. The recombinant proteins are then either secreted into the surrounding media or into the perinuclear space, or made intracellularly and extracted from the cells. A number of well-known technologies are utilized for purifying desired proteins from their source material. Purification processes include procedures in which the protein of interest is separated from the source materials on the basis of solubility, ionic charge, molecular size, adsorption properties, and specific binding to other molecules.
When developing processes for commercial production of therapeutically important proteins, purification of the protein sample to remove undesired species in an efficient manner is highly desirable. The present invention provides a purification method for removal of high molecular weight species from a protein sample using two sequential chromatographic steps.